• Journal of Animal Science and Technology
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• v.53 no.1
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• pp.15-22
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• 2011

The purpose of this study was to analyse of association between growth traits and single nucleotide polymorphisms (SNPs) polymorphism of phosphoglycerate kinase 2 (PGK 2) gene in pigs. The birth weight of piglet influences on weaning weight and survival rate that are import economic traits in pig industry. Also, these growth traits are representative factor to decrease a period getting to marketing weight as well as growth rate in pig. The PGK 2 is an isozyme that catalyzes the first ATP-generating step in the glycolytic pathwayand important enzyme related with energy metabolism. Twenty of SNPs were discoveredby genome structure analysis that compares the sequence on promoter and transcription region of PGK 2 gene in porcine chromosome 7. An association between PGK 2 SNPs and growth traits was analyzed in $F_2$ reciprocal-crossbred population between korean native pig (KNP) and Landrace. Association analysis indicated that polymorphism of the PGK 2 gene promoter region has significant effects on weight at birth (p<0.01) and weight at 3 weeks of age (p<0.0001). These results suggest that PGK 2 gene polymorphism was associated with energy metabolism and physiological function of growth in pig.

• Microbiology and Biotechnology Letters
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• v.35 no.3
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• pp.184-190
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• 2007

Approximately 2.0 kb of the promoter region of the Pichia pastoris phosphoglycerate kinase gene (PGK1) was reduced to a 266 bp fragment and this minimized portion was used for construction of a new episomal constitutive expression vector in P. pastoris. As an approach to developing a constitutive expression vector in P. pastoris, the GAP promoter region of the Pichia expression vector pGAPZB was replaced with sequentially deleted PGK1 promoter fragments fused to a beta-galactosidase gene. When a lacZ gene was used as a reporter gene, PGK1 promoter strength was lower than that of the constitutive GAP promoter but it was higher than TEF1. We report here the development of the pPGKZ-E vector as a new episomal expression vector for heterologous gene expression by removing non-essential regions of the PGK1 promoter. This broadens the choice of episomal expression vectors for controlled constitutive expression in P. pastoris.

• Journal of Life Science
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• v.23 no.2
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• pp.190-196
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• 2013

The sea squirt, Halocynthia roretzi, has experienced mass mortality due to softness syndrome. The identification of disease-induced genes can provide insights into the development of this syndrome. To identify the genes, we performed differentially expressed gene (DEG) analysis. The expression of the phosphoglycerate kinase (HrPGK) gene was significantly decreased in diseased sea squirts compared to normal ones. We confirmed the result of the DEG analysis through RT-PCR and real-time PCR. In addition, we detected single nucleotide polymorphisms at position -106 (A/T) and -254 (G/T) in the HrPGK gene promoter by genotyping analysis. At the -106 site of the HrPGK gene, the frequency of the AA allele in disease-resistant sea squirts was about two-fold higher than that of sensitive ones, and the frequency of the TT allele in the disease-resistant sea squirts was about six-fold lower. At the -254 site of the HrPGK gene, the frequency of the GT and the GG allele was approximately two-fold higher and two-fold lower, respectively, in the disease-resistant sea squirts compared to the disease-sensitive ones. Analysis of the relationship between the genotypic variation at the -106/-254 promoter and the expression of HrPGK mRNA showed that HrPGK mRNA expression was higher in the -106/-254 AA/GT genotype samples than in the -106/254 TT/GG genotype ones. These results show that sea squirts harboring the AA/GT genotype may have more resistance to mortality than the sea squirts with other genotypes.

• Microbiology and Biotechnology Letters
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• v.29 no.4
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• pp.206-211
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• 2001

In order to overexpress constitutively the Kluyveromyces marxianus exoinulinase gene (INUI) in Saccharomyces cerevisiae, four episomal expression systems employing GAPDH, ADHI, PGK and ENOI promoters were constructed as p YIGP aADHI -INU, pPGK-INU, and pENOI- INU plasmids respectively, When S cereviais transformants harboring each plasmid were batchwisely cultivated in the fermentor containing 5% glucose medium no significant differences in the cell growth are observed How- ever the experession level of exoinulinase and plasmid stability showed a strong dependency on the promoter employed. The expression levels of exoinulinase were about 1.70 unit/ml for GAPDH promoter 1.67 unit/ml for PGK promoter, 1.29 unit /ml for ADH1 promoter, and 0.80 unit/ml for ENOl promoter. The plasmid stabilites were maintaines above 80% in all experession systems. except the GAPDH promoter system of 55%, Based on the plas- mid stability and expression level of exoinulinase the ADHl and PGK promoter system were selected for the fed - batch culture to overproduce exoinulinase By the intermittent feeding of yeast extract and glucose, both promoter systems gave the cell concentration of about 30 g-dry cell weight/1 byt the maximal exoinulinase activity of 3.70 unit/ml and plasmid stability of 96% in the ADH1 promoter were higher than those (2.70 unit/ml, 80%) of PGK sys- tem Taking into account the plasmid stability and extended culture time the ADH1 promoter systems would be the most feasible expression systems for the constitutive overproduction of exoinulinase through high cell-density fed- batch cultures using non-selective rich medium.

• BMB Reports
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• v.29 no.4
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• pp.359-364
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• 1996

By both in vitro hydroxylamine mutagenesis of the wild type 3-phosphoglycerate kinase gene (PGK) promoter DNA and insertion of the leu2-d gene, we have created yeast expression vectors potentially useful for production of eukaryotic genes in yeast. The guanine (G) to adenine (A) change at the -3 position from the ATG start codon of the PGK promoter-based vector rendered a 6~7 times elevated expression of the adjacent eukaryotic gene, and insertion of the leu2-d gene in the vector containing the mutated PGK promoter further enhanced the expression of the gene. When expression of the AIDS virus HIV1-gagP17 gene in a lacZ fusion form was examined with this new vector, a 15 times higher level of expression than that from the original PGK promoter was observed. Northern and Southern analysis showed that this elevated expression is due to the production of a high copy number of mRNA by leu2-d gene functioning and by efficient translation of the produced mRNA. Thus, the vector that contained the A at the -3 position from the ATG start codon in the promoter region and the leu2-d gene shows increased expression capability and will be potentially useful for production of eukaryotic genes in yeast.

• Reproductive and Developmental Biology
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• v.33 no.1
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• pp.63-69
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• 2009

In this study we tried to construct a more efficient tetracycline-inducible gene expression system by replacing previous key elements with more advance ones. At the beginning, we substituted PGK (phophoglycerate kinase) promoter for CMV (cytomegalovirus) promoter to control "$rtTA2^sM2$" which has been known for high induction efficiency in response to tetracycline. With this modification, expression of the EGFP marker gene under the induction condition was significantly increased. Next, we replaced "TRE" fragment with a modified version named "TRE-tighf" which has been reported to have higher affinity and specificity to the transactivator by minor base change of the "TRE" DNA fragment sequence. Use of "TRE-tighf" instead of "TRE" resulted in more than 10 fold increment in terms of induction efficiency and significant decrement of background expression in non-inducible condition. By combining PGK promoter and "TRE-tight" fragment, we could upgrade previous tetracycline-inducible system to show more stringent turn on/off gene switch ability and stronger expression of the gene of our interest. Use of this newly developed system must be very helpful to the studies of gene expression, especially to the transgenic animal study in which non-controllable constitutive expression of the transgene has been one of the urgent problems to be solved.

• The Korean Journal of Zoology
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• v.39 no.1
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• pp.75-88
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• 1996

OTF9-63 (OTF9) and P19S1O1A1 (P19) embryonal carcinoma (EC) cells were examined for their ability to produce the readivation of inactive X chromosomes from somatic cells. They were hybridized with various somatic cells and resulting HATr EC-somatic cell clones were analysed for their morphology, chromosomal replication pafterns and expression proffies of X-linked and distantiy located genes, Hprt and Pgk-1. The results demonstrated that 0RF9 cells could reactivate the inactive X chromosome whereas P19 cells could not. In adition, EC-somatic cell hybrids tended to reduce the number of sex chromosomes in long-term culture, resulting m 1:2 ratio of sex chromosomes to autosomes The use of EC cell hybrids provides an experimental system for studying the mechanism(s) of the X-reactivatio that is initiated and maintained from meiotic prophase of oogenesis to early embryogenesis.

• Korean Journal of Environmental Biology
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• v.22 no.3
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• pp.472-477
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• 2004

To study the radiation related gene expression in mutants of Bacillus lentimorbus WJ5 induced by gamm radiation, the simultaneous gene expression was analyzed by DNA micro array. We constructed DNA chips including two thousand randomly digested genome spots of B. lentimorbus WJ5 and compared its quantitative aspect with seven mutants induced by gamma radiation $(^{60}/Co)$. From the cluster analysis of gene expression pattern, totally 408 genes were expressed and 27 genes were significantly upregulated by the gamma radiation in all mutants. Especially, genes involved in repair (mutL, mutM), energy metabolism (acsA, sdhB, pgk, yhjB, citB), protease (npr), and reduction response to oxidative stress (HMM) were simultaneously upregulated. It seems that the induction of the direct and/or indirect repair related genes in mutants induced by gamma radiation could be remarkably different from the adaptive responses against acute exposure to radiation.

• Journal of Embryo Transfer
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• v.22 no.3
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• pp.179-184
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• 2007

Culture method of somatic cells is one of the important factors in the production of transgenic pigs by somatic cell nuclear transfer. In this study, we established an efficient culture method of porcine fetal fibroblasts. Porcine fetal fibroblasts were isolated from 33-day-old fetuses. The proliferation of porcine fetal fibroblasts was analyzed by different serum types and culture media. The cultures in medium supplied 15% ES screened FBS showed faster increase in cell number than 15% FBS. Also, fetal fibroblasts have been propagated continuously for $7{\sim}8$ passages in ES modified DMEM and DMEM medium. We transfected $PGK-neo^r$ vector (pKJ2) into porcine fetal fibroblasts to estimate colony formation in this culture condition. The formation of colonies was confirmed in the medium containing $300\;{\mu}g/ml$ G418 at 12 day. These data show that this culture system can be used screening of porcine somatic cells transfected transgene.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.11a
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• pp.147-172
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• 1994

Aziridinylbenzoquinones such as 3, 6-diaziridinyl-1, 4-benzoquinone (DZQ) and its 2, 5-methyl analog (MeDZQ) require bioreductive activation in order to elicit their anticancer activities. To determine the involvement of DTD in the activation of these drugs, we have used a ligation-mediated polymerase chain reaction to map the intracellular alkylation sites in a sing1e copy gene at the nucleotide level. We have performed this analysis in two human colon carcinoma cells, one proficient (HT-29) and one deficient (BE) in DT-diaphorase (DTD) activity. In the DTD proficient HT-29 cell line, DZQ and MeDZQ were found to alkylate both 5'-(A/T)G(C)-3' and 5'-(A/T)A-3' sequences. This is consistent with the nucleotide preferences observed when DZQ and MeDZQ are activated by purified DTD to reactive metabolites capable of alkylating DNA in vitro [Lee, C. -S., Hartley, J. A., Berardini, M. D., Butler, J., Siegel., D., Ross, D., & Gibson, N. W. (1992) Biochemistry, 31: 3019-3025]. Surprisingly in the DTD-deficient BE cell line a pattern of alkylation induced by DZQ and MeDZQ similar to that observed in the DTD-proficient HT-29 cells was observed. This suggests that reductive enzymes other than DTD can be involved in activating DZQ and MeDZQ to DNA reactive species in vivo.